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    <title>Assessing Quantitative Accuracy</title>
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        <h1 align="center">Assessing Quantitative Accuracy</h1>
		<p align="left">To provide a example of how quantitative 
		accuracy can be evaluated, data has been taken from the Experiment Demo 
		Database is used to demonstrate two central aspects, which are assay 
		precision and absolute accuracy. </p>
		<p align="left">To assess the general level of assay 
		precision, target quantities generated from the three runs for the 12240, 46630 
		and GAPDH targets within the COL 1 sample are compared:</p>
		<p align="center">
		<img border="0" src="images/lre_quantification.gif" width="262" height="153"></p>
		<p align="left">The CV values provide a general indication of run to run 
		variance, producing an average of ±7.5%. This is reflective of the 
		high level of precision that can be achieved when a high performance 
		instrument (AB 7500) is combined with high performance amplicons. </p>
		<p align="left">However, this type of analysis cannot be used to assess 
		absolute accuracy. As described in the <a href="lda_overview.html">LDA Overview</a>, Poisson 
		distribution can be used to conduct absolute quantification that is 
		independent of real-time PCR, accomplished by diluting the target to below one 
		molecule per reaction and conducting moderately large number of replicate reactions (16-32). Based on the number of nil reactions (i.e. 
		those that fail to produce an amplification profile), target 
		quantity can be determined using the equation:</p>
		<p align="center">
		<img border="0" src="images/lda_equation.gif" width="186" height="71"></p>
		<p align="left">where &quot;#nil&quot; is the number of amplification reactions 
		that fail to produce an amplification profile (i.e. 0 N aliquots), &quot;#total&quot; is the total 
		number of replicate reactions conducted and &quot;N<sub>av</sub>&quot; is the 
		target concentration. </p>
		<p align="center">
		<img border="0" src="images/lda_quantification.gif" width="294" height="167"></p>
		<p align="left">An important qualification is that reactions that 
		produce non-specific amplification products must either be absent or can 
		be identified, as such reactions generate false positives. Indeed, the GAPDH amplicon used in 
		this analysis did not fulfill this requirement, as it generated a large 
		number of small, non-specific products. While this is generally not 
		typical, it does provide an example of how LDA can also provide useful 
		insights into amplicon performance that may not be evident with high target quantities. </p>
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